Kallikrein is a protease which is widely distributed in plasma and tissues of various animals including human beings. An enzymatic system known as the kallikrein-kinin system acts in vivo. It has a close relationship with various other enzymatic reaction systems such as a renin-angiotensin system, a blood clotting system, a fibrinolysis system, a complement system as well as catecholamine and arachidonic acid cascade mainly related to prostaglandins, leukotrienes and thromboxanes. Accordingly, the kallikrein-kinin system is closely associated with blood pressure regulating action and blood clotting-fibrinolysis-complement system action. Bioregulation and an improving action for peripheral circulation by various physiologically active substances produced by an arachidonic acid cascade are also related to the kallikrein-kinin system. The kallikrein-kinin system plays an important role in the regulation of functions in vivo.
Kinins, such as bradykinin, are produced in the kallikrein-kinin system. They exhibit various physiological actions such as a decrease in blood pressure due to dilation of peripheral blood vessels, promotion of permeability of blood vessels, contraction or relaxation of smooth muscle, induction of pain, induction of inflammation, migration of leucocytes, liberation of catecholamine from the adrenal cortex, etc. Kinins are also known as mediators in acute inflammations including allergic reactions whereby their existence in vivo has a profound significance.
With respect to this plasma kallikrein-kinin system, it has been believed that a blood coagulation factor XII (a Hageman factor, hereinafter abbreviated as FXII) is activated in vivo by injury and invasive stimulation to tissues whereby a series of enzymatic reaction systems results. Thus, as shown in FIG. 1, the activated blood coagulation factor XII or "Activated FXII" (hereinafter, abbreviated as FXIIa) acts on the plasma prekallikrein which exists in the same plasma to convert it to a plasma kallikrein which is an enzyme in an activated form. Then the plasma kallikrein acts on a high-molecular-weight kininogen (hereinafter abbreviated as an HK) to liberate bradykinin.
Kinins such as bradykinin which are liberated by the enzymatic reaction of the kallikrein-kinin system exhibit various physiological actions as mentioned already. Accordingly, substances which inhibit the action of bradykinin or substances which inhibit the production of bradykinin by interfering with the reactions in the plasma kallikrein-kinin system may be useful as antiinflammatory, analgesic and antiallergic agents.
Further, FXIIa is an important factor in the initiating stage of the intrinsic blood clotting system and the fibrolysis system. Thus, substances which affect the production of FXIIa may be useful as drugs in the areas of blood clotting and fibrinolysis.
Therefore, establishment of a method for measuring the degree to which substances or compounds or components inhibit or promote the reactions in the plasma kallikrein-kinin system in a simple, easy, quick and precise manner is a very important means for ascertaining the action which helps regulation of the above-mentioned bioregulation systems. It is also useful for developing drugs for regulating or controlling the bioregulation systems.
When screenings or the like of drugs using the plasma kallikrein-kinin system are carried out in vitrO, activation of FXII by an invasive stimulation to tissues and injury such as an intravital reaction cannot be conducted. A substance which activates the FXII may be added to an isolated plasma to carry out a reaction which induces a plasma kallikrein-kinin system reaction in vitro.
However, the plasma of animals contain various components in addition to the above-mentioned components. For example, components which have an effect (such as an inhibition or a promotion) on the plasma kallikrein-kinin reaction system and other unknown factors are contained in animal plasma. Accordingly, measuring the activity of the test substance towards the production of the physiologically active substances using animal plasma per se to screen drugs or the like is complicated. The various factors containing unknown components which are in the animal plasma per se may affect the plasma kallikrein-kinin reaction system. Consequently, controlling the reaction system is highly technical and complex when animal plasma per se is used as a source of reactants.
The present invention provides a method for measuring the physiological action of the tested substance in an easy, simple, prompt and precise manner. Factors containing unknown components which may affect the plasma kallikrein-kinin reaction system are eliminated by the use of a reconstituted plasma kallikrein-kinin reaction system. The drug screening reactions may be carried out in vitro in the reconstituted plasma kallikrein-kinin reaction system. The reconstituted plasma kallikrein-kinin system may be obtained by combining components isolated from the plasma kallikrein-kinin system.